Project Summary: Colorectal cancer conveys a high mortality risk as the second most common cause of cancer in the US. While newer chemotherapy regimens have improved survival, 37% of all patients diagnosed with CRC in the United States will die of this disease. Clearly, new tumor targets are needed to develop improved treatment approaches. Harnessing natural anti-tumor immunity is a promising approach to develop new therapies. develop new therapies. We have found that one potential way to do this is by blocking a critical cytokine that induces tumor cell proliferation and invasion along with inhibitory immune responses in CRC. Granulocyte colony-stimulating factor (G-CSF) is a key cytokine present in 88% of human colorectal tumors. Blockade of this cytokine in a mouse model of colorectal cancer led to activation of protective immune responses and neoplasm regression. G-CSF may induce regulatory T cell accumulation and potent inhibition of cytotoxic cell responses. Since these responses may have critical tumor promoting functions, we hypothesize that G-CSF blockade is protective in CRC by inhibiting tumor progression and inducing anti-tumor immunity. To test this hypothesis, the following Specific Aims will be completed: Specific Aim 1: Delineate the role of G-CSF/G-CSFR in colorectal cancer progression and the potential of blockade as a therapeutic approach. We hypothesize that G-CSF/G-CSFR inhibition will prevent or regress colorectal cancer metastasis. This will be examined by knocking down G-CSF/G-CSFR in tumor cells, overexpressing G-CSF/G-CSFR in tumor cells, and employing therapeutic approaches in mouse models of CRC metastasis. Human CRC tissues will be examined for an association with G-CSF/G-CSFR expression and metastasis. Specific Aim 2: Delineate the role of G-CSF/G-CSFR inhibition on enhancing anti-tumor immunity to protect against CRC progression. We hypothesize that G-CSF induces a tumor immune evasive microenvironment by inducing immune cell IL-10 production and by inhibiting cytotoxic immune cell responses. The direct effects of G-CSF on myeloid cell and T cell IL-10 production will be examined. The direct effects of G-CSF on NK and CD8+ T cells will be examined along with indirect effects through modulation of IL-10 in the tumor microenvironment will be examined. The ability of G-CSF to inhibit NK cell and CD8+ T cell tumor lytic function will be assessed. Human CRC and liver metastasis tissues with high vs low G-CSF/G-CSFR expression will be examined for NK and CD8+ T cell activity. Thus, for this project, we will examine multiple mechanisms of G-CSF blockade on tumor proliferation and invasion, activation of protective myeloid and T cell responses, and test blockade of this pathway in an invasive colorectal cancer model and in human tumor tissues in translational approaches that could lead to a new treatment for colorectal cancer.